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Abstract

DEVELOPMENT AND VALIDATION OF LC-MS/MS METHOD FOR SIMULTANEOUS QUANTIFICATION OF METFORMIN AND NATEGLINIDE IN HUMAN PLASMA AND ITS APPLICATION TO A PHARMACOKINETIC STUDY

Raja Haranadha Babu Chunduri* and Gowri Sankar Dannana

ABSTRACT

A selective, sensitive and rapid LC-MS/MS method has been developed and validated for simultaneous quantification of metformin and nateglinide in human plasma using repaglinide as internal standard (IS). The analytes and IS were extracted from 100 μL of plasma by liquid-liquid extraction technique using methyl tert - butyl ether which offers high sensitivity, wide linearity without interferences form endogenous matrix components. Chromatographic separation was achieved in 1.50 min run time on Hypersil gold C18 column using a 5 mM ammonium formate/acetonitrile mobile phase in gradient mode. The quantification of target compounds was performed in a positive electrospray ionization mode and multiple reaction monitoring (MRM). The proposed method was validated over the concentration ranges of 1-20000 ng/mL for metformin and 0.01-200 ng/mL for nateglinide. The intra- and inter-day precision and accuracy results were acceptable as per FDA guidelines. Stability of compounds were established in a battery of stability studies, i.e. bench top, auto sampler, dry extract and long term storage stability as well as freeze-thaw cycles. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic studies.

Keywords: Metformin; Nateglinide; LC-MS/MS.


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